The transition from a functional photosynthetic organ to an actively degenerating and nutrient-recycling tissue in a leaf's life history represents the onset of leaf senescence. This onset is a developmental switch that involves dramatic differential gene expression. Differential gene expression is believed to play an important role in leaf senescence. In a senescing leaf, many genes that are expressed in green leaves, including those genes involved in photosynthesis, are down-regulated, while a subset of genes, generally referred to as senescence-associated genes (SAGs), are up-regulated. Leaf senescence is under direct nuclear control, and SAG expression is required for senescence to proceed. Inhibitors of transcription or translation prevent leaves from senescing (Buchanan-Wollaston et al., “The Molecular Analysis of Leaf Senescence—A Genomics Approach,” Plant Biotechnology Journal 1:3-22 (2003); Guo et al., “Leaf Senescence: Signals, Execution, and Regulation,” Current Topics in Developmental Biology 71:82-112 (2005); Hadfield et al., “Programmed Senescence of Plant Organs,” Cell Death Differ. 4:662-670 (1997); Lim et al., “The Molecular and Genetic Control of Leaf Senescence and Longevity in Arabidopsis,” Current Topics in Developmental Biology 67:49-83 (2005); Smart, “Gene Expression During Leaf Senescence,” New Phytologist 126:419-448 (1994)). For the past decade, much effort has been made to isolate SAGs, and hundreds of SAGs have been cloned from various plant species including Arabidopsis, barley, Brassica, maize, cucumber, rice, tobacco, radish, asparagus and soybean (Buchanan-Wollaston et al., “The Molecular Analysis of Leaf Senescence—A Genomics Approach,” Plant Biotechnology Journal 1:3-22 (2003); Gepstein et al., “Large-Scale Identification of Leaf Senescence-Associated Genes,” Plant Journal 36:629-642 (2003); He et al., “Molecular Characteristics of Leaf Senescence,” In Recent Research Developments in Plant Molecular Biology, Kerala, India: Research Signpost, pp. 1-17 (2003)). Recent application of genomics approaches has led to the identification of thousands of potential SAGs (Andersson et al., “A Transcriptional Timetable of Autumn Senescence,” Genome Biology 5:R24 (2004); Bhalerao et al., “Gene Expression in Autumn Leaves,” Plant Physiology 131:430-442 (2003); Buchanan-Wollaston et al., “The Molecular Analysis of Leaf Senescence—A Genomics Approach,” Plant Biotechnology Journal 1:3-22 (2003); Buchanan-Wollaston et al., “Comparative Transcriptome Analysis Reveals Significant Differences in Gene Expression and Signalling Pathways Between Developmental and Dark/Starvation-Induced Senescence in Arabidopsis,” The Plant Journal 42:567-585 (2005); Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004); Lin et al., “Molecular Events in Senescing Arabidopsis Leaves,” Plant Journal 39:612-628 (2004); Zentgraf et al., “Senescence—Related Gene Expression Profiles of Rosette Leaves of Arabidopsis Thaliana: Leaf Age Versus Plant Age,” Plant Biology 6:178-183 (2004)). Analysis of a leaf senescence EST database (dbEST) indicated that approximately 10% (approximately 2500) of the Arabidopsis genes are expressed in senescent leaves (Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004)). Microarray analysis of the global gene expression changes during developmental leaf senescence in Arabidopsis has led to the identification of more than 800 genes that show a reproducible increase in transcript abundance (Buchanan-Wollaston et al., “Comparative Transcriptome Analysis Reveals Significant Differences in Gene Expression and Signalling Pathways Between Developmental and Dark/Starvation-Induced Senescence in Arabidopsis,” The Plant Journal 42:567-585 (2005)).
Changes of gene expression are often regulated by transcription factors that bind to specific cis elements of target gene promoters, resulting in the activation and/or suppression of the target genes. There are approximately 1500 transcription factor genes in the Arabidopsis genome that belong to more than 30 gene families based on their DNA-binding domains (Riechmann et al., “Arabidopsis Transcription Factors: Genome-Wide Comparative Analysis Among Eukaryotes,” Science 290:2105-2110 (2000)). Microarray analysis has identified 96 transcription factor genes with at least a threefold upregulation during leaf senescence (Buchanan-Wollaston et al., “Comparative Transcriptome Analysis Reveals Significant Differences in Gene Expression and Signalling Pathways Between Developmental and Dark/Starvation-Induced Senescence in Arabidopsis,” The Plant Journal 42:567-585 (2005)), and analysis of the leaf senescence dbEST revealed 134 unique genes that encode transcription factors representing 20 different gene families (Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004)). Among the largest transcription factor groups are NAC, WRKY, C2H2 type zinc finger, AP2/EREBP, and MYB proteins (Buchanan-Wollaston et al., “Comparative Transcriptome Analysis Reveals Significant Differences in Gene Expression and Signalling Pathways Between Developmental and Dark/Starvation-Induced Senescence in Arabidopsis,” The Plant Journal 42:567-585 (2005); Chen et al., “Expression Profile Matrix of Arabidopsis Transcription Factor Genes Suggests Their Putative Functions in Response to Environmental Stresses,” Plant Cell 14:559-574 (2002); Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004); Lin et al., “Molecular Events in Senescing Arabidopsis Leaves,” Plant Journal 39:612-628 (2004)). Two WRKY transcription factor genes have been studied: WRKY53 plays an important role in controlling leaf senescence (Hinderhofer et al., “Identification of a Transcription Factor Specifically Expressed at the Onset of Leaf Senescence,” Planta 213:469-473 (2001); Miao et al., “Targets of the WRKY53 Transcription Factor and Its Role During Leaf Senescence in Arabidopsis,” Plant Mol Biol 55:853-867 (2004); Robatzek et al., “Targets of AtWRKY6 Regulation During Plant Senescence and Pathogen Defense,” Genes Dev 16:1139-1149 (2002)), while suppression of WRKY6 expression has little effect on either the onset or the progression of leaf senescence (Hinderhofer et al., “Identification of a Transcription Factor Specifically Expressed at the Onset of Leaf Senescence,” Planta 213:469-473 (2001); Miao et al., “Targets of the WRKY53 Transcription Factor and Its Role During Leaf Senescence in Arabidopsis,” Plant Mol Biol 55:853-867 (2004); Robatzek et al., “Targets of AtWRKY6 Regulation During Plant Senescence and Pathogen Defense,” Genes Dev 16:1139-1149 (2002)). The potential functions of the majority of the leaf senescence-associated transcription factors remain to be elucidated.
A total of 20 genes encoding NAC transcription factors are in the leaf senescence dbEST (Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004)), representing almost one-fifth of all the predicted 109 members of the NAC superfamily in Arabidopsis (Riechmann et al., “Arabidopsis Transcription Factors: Genome-Wide Comparative Analysis Among Eukaryotes,” Science 290:2105-2110 (2000)). The NAC domain was originally defined by the highly conserved N-termini of the petunia NAM (NO APICAL MERISTEM) and Arabidopsis ATAF1 and CUC2 (CUP-SHAPED COTYLEDON2) genes. It exists widely in plants but not in other eukaryotes. Roles of the NAC family genes include embryo and shoot meristem development, lateral root formation, auxin signaling, defense, and abiotic stress response (Olsen et al., “NAC Transcription Factors: Structurally Distinct, Functionally Diverse,” Trends Plant Sci 10:79-87 (2005)). Expression of the NAC family genes in senescing leaves has been reported by several groups (Andersson et al., “A Transcriptional Timetable of Autumn Senescence,” Genome Biology 5 (2004); Buchanan-Wollaston et al., “Comparative Transcriptome Analysis Reveals Significant Differences in Gene Expression and Signalling Pathways Between Developmental and Dark/Starvation-Induced Senescence in Arabidopsis,” The Plant Journal 42:567-585 (2005); Guo et al., “Transcriptome of Arabidopsis Leaf Senescence,” Plant Cell and Environment 27:521-549 (2004); John et al., “Cloning and Characterization of Tomato Leaf Senescence-Related cDNAs,” Plant Molecular Biology 33:641-651 (1997); Lin et al., “Molecular Events in Senescing Arabidopsis Leaves,” Plant Journal 39:612-628 (2004)), but whether these genes play a part in leaf senescence is unknown.
The present invention is directed to overcoming these deficiencies in the art.